To assess the potential role of ras oncogene activation and p53, p16/CDKN2 tumor suppressor gene mutations in the development of human prostate carcinoma, nine cases of histologically heterogeneous prostate tumors obtained from total prostatectomies were probed for these specific events. Each tumors was divided into 5-10 areas according to different growth or hostologic patterns. Targeted DNA sequences coding for ras, p53 and p16/CDKN2 were amplified using the polymerase chain reaction and analyzed by single-strand conformational polymorphisms, and confirmed by direct DNA sequencing. Point mutations of the ras gene were found in 3 of the 9 tumors. Two cotained K-ras codon 13 and H-ras codon 61 mutations, found in only one and three areas of each lesion, respectively. The third tumor contained 2 different point mutations in K-ras codons 13 and 61 in different foci of sample. Loss of heterozygosity (LOH) at the polymorphic codon 72 in the p53 gene was detected in 2 of 4 informative cases (50%) showing fragment cleavage by restriction fragment length polymorphism analysis. Mutations in p53, missense transversions, single base insertions and two base deletions, were also detected in three tumors. No homozygous deletions of p16/CDKN2 gene were observed in any of the carcinomas, but two of the nine tumors demonstrated missense mutations in exon 2 of p16/CDKN2 gene. The mutations were detected in only one and two foci, respectively, out of six and ten selected tumor areas. The present results reveal mutated ras, p53 and p16/CDKN2 occasionally occurring in small foci of the tumor, and that genetic mutations in p53, as opposed to those in ras and p16/CDKN2 are more closely associated with invasive growth of heterogeneous prostate carcinoma.